Our goal is to develop a systematic approach to study genetic correction of the defect in cystic fibrosis lung disease. Specific goals include the following: First, retrovirus vectors containing the normal cDNA for the CFTR gene will be constructed and tested for their ability to mediate gene transfer into both human primary epithelial cells and CF cell lines. The stability of CFTR in these vectors will be evaluated. We will take advantage of preliminary results that have suggested specific vectors to use for efficient expression in epithelial cells. In addition, several approaches are proposed to increase the efficiency of retrovirus vector gene transfer, including the novel use of bicistronic vectors. A second goal is to investigate the airway biology pertinent to the introduction of foreign genes by retrovirus vectors. Primary human epithelium and immortalized cell lines will be used to study the efficiency of retrovirus gene transfer as well as the fate of vector expression during differentiation of airway epithelium. Also,, -.the types of cells that serve as targets for retrovirus vector infection will be determined. Selected experiments using ferrets will be used to study issues presently impossible to study in humans including: the correlation of retrovirus vector infection in vitro and in vivo; the persistence of vector expression in vivo; the effect of maturational factors on vector infection; and the effect of limited airway damage on the efficiency of retrovirus vector gene transfer. A third goal is to determine the fraction of CF airway cells in which the normal CFTR needs to be expressed in order for affected epithelia to attain normal bioelectric properties.